•296 A Bacterial Disease of Turnip (Brassica Xapus) 



by iminediatelv placing the petri-dishes in an incubator at GO '. LoefHer's 

 method of staining was first employed \vithout success, but with this 

 method, the organisms showed uniform staining, except at one pole 

 which was somewhat hyaline. When however the mordant was made 

 sUghtly alkahne with caustic-soda (J c.c. of 1 per cent, alkali per 10 c.c. 

 of tannin) most of the shdes after staining showed the presence of a 

 single long polar Hagellum. Van Ermengen's stain, pre])ared according 

 to the usual formula, also showed the presence of the cihum. No varia- 

 tion in the number or .situation of tlie flagella was seen. 



Infection Experiment.s. 



The next step was to establisli the disease in healthy plants. It was 

 decided to carry out infection of healthy plants by employing cultures 

 from inoculated blocks. Preparations were accordingly made for inocu- 

 lating a series of blocks from young cultures taken from the agar-streak 

 tubes that had last been employed in the flagella staining. The latter 

 process had entailed repeated failures and fresh attempts always involved 

 the use of fresh subcultures. It was reckoned that the tubes now em- 

 ployed for inoculating the turnip blocks contained cultures which had 

 passed through some 20 generations over the medium. After the usual 

 period of incubation, the inoculated blocks showed little or no signs of 

 disease. Some of the blocks showed a slight browning around the 

 inoculated part and made no further advance; others did not appear 

 to have taken the disease at all. It seemed that the organism had either 

 lost its virulence in its repeated passages through the medium, or that 

 the turnip, through prolonged cultivation had passed over into a state 

 of resistance to attack. It was therefore decided to submit sterile blocks 

 of turnip to special treatment on the lines followed by Laurent for 

 rendering resistant tubers sensitive to bacterial attack, by immersing 

 them in alkaline solutions. Accordingly the blocks were first soaked for 

 an hour, some in 0-25 per cent., some in 0-.5 per cent, and others in 

 1 per cent, caustic-soda solutions. They were then transferred to sterile 

 test-tubes and inoculated from the same agar-streak cultures that had 

 been employed on the supposed resistant blocks. After prolonged incuba- 

 tion the parasite made no more progress on the blocks thus treated than 

 on the control ones which had only been soaked in water previous to 

 inoculation. It was therefore concluded that the organism had lost 

 virulence by so many passages through the artificial medium. When, 

 however, another series of turnip blocks was inoculated from an agar- 



